The present invention relates to apparatus for dispensing a sample for subsequent analysis by mass spectrometry, and to a method of fabricating such apparatus.
Mass spectrometry is a performant analytical technique, which provides information about the chemical nature of analyzed molecules. The application domain of mass spectrometry analysis has grown in the past few years with the development of technologies which allow the injection of a broad range of molecules. Among the different feeding processes, the electrospray or nanospray is a method of choice, where the sample is diluted in a solvent and sprayed from a flow of solution under a high voltage ionization.
Recent developments have allowed the automation of the feeding of samples to a mass spectrometer (MS) in order to enhance the throughput of the analysis. Indeed, mass spectrometry analysis is a rapid technique that can work with a scan rate of more that 10 kHz. On the other hand, more and more performant informatic tools allow the rapid treatment of data for example with new software that is able to compare the obtained data with a full database in a very short time. Therefore, the need for fast and automated sampling systems is growing.
Different designs have been presented for fabricating a nanospray. The more common way is to pull a capillary and to metallize its external surface. The capillary is then connected to a nanospray tip and placed in front of a mass spectrometer. This method is convenient but difficult to fabricate in a reproducible way. Such a system is however easier to fabricate in a planar microchip, also called a microfluidic device as used in the present invention.
A standard electrospray is composed of a capillary end that is surrounded by a metallic substrate to apply the high voltage, while sheath liquid and sheath flow are provided to enhance the performances of the ionization and evaporation of the solvent. More recently, various approaches have been investigated to provide sheathless electrospray systems where a capillary with small enough dimensions insures an efficient spray (see Wahl J H et al, Electrophoresis, 1993, vol. 14, p 448). Alternatively, WO 98/35226 describes inserting a platinum wire inside the capillary and gluing it with epoxy in order to apply the high voltage at the exit of the capillary.
Microfluidic devices are composed of a plate or a film with covered microchannel networks and are principally developed for electrophoretic separations. These systems are coupled to a mass spectrometer in an ever-growing number of cases. Such a coupling is ensured by a nanospray interface, which is able to work with a slow flow rate (typically 1 to 500 nL/min). Several approaches have been shown to reach this goal, mainly glass chips being coupled to an MS through a liquid junction (Figues et al, Analytical Chemistry, 1998 vol. 70 p 3728) or by adjusting a nano-tip at the end of the microchip (Li et al, Analytical Chemistry, 1999 vol. 71 p 3065). In an alternative system described by Ramsey et al, Analytical Chemistry, 1997, vol. 69 p 1174, the microchannel end has been directly coupled to an MS, by applying the potential from a side capillary. An interface between a polymer-based microchip in PDMS and a capillary for the nanospray coupling has also been suggested by Chan et al, Analytical Chemistry, 1999 vol. 71 p 4437. An interesting point is that this configuration allows the multiplexing of capillaries and opens the way to multiple analyses in a minimum of time. In another concept described by Xu et al, Analytical Chemistry, 1998 vol. 70 p 17, the solution is pumped through a membrane cut-off and the solution is driven to a metallized spray tip to the MS. In WO 98/35376, an electrospray nozzle has a filter structure that is integrated between the sample inlet and the channel tip outlet. The concept of multiplexing a nanospray is presented in U.S. Pat. No. 5,872,010, in which a covered microchannel array is used for feeding the mass spectrometer. The high voltage is applied in a sample or buffer reservoir and not by a conductive region inside the covered microchannels. Numerous methods have been described in which microchips have been coupled to an MS, through more or less direct interfaces.
In another format, a polycarbonate chip was fabricated by mechanically machining a cone in a polymer block, and using a laser to etch a capillary system in one of the polymer blocks that ended in an electrospray nozzle. (Wen et al. Electrophoresis 2000, 21, 191–197). This format necessitates the etching of a polymer by machining in order to remove material to form a tip shape more suited for electrospraying.
In another work, Kim et al. (Kim et al. Electrophoresis 2001, 22, 3993–3999) presented a way of fabricating arrays of electrospray in PDMS by casting that can be directly interfaced to the MS. Nevertheless, because of the fairly thick polymer they use for stability, they have to assist the flow with nitrogen in order to generate a stable spray. Otherwise, some authors (WO 00/15321) presented a way of fabricating nanospray nozzles in silicone that can be further interfaced with glass or plastic chips. Nevertheless, in this work, the authors did not mention machining a plastics chip to form the nozzle.
The integration of microelectrodes, microholes and microionodes (i.e. composite ion permeable membranes) has been achieved recently by photoablation or by plasma etching, for the electrochemical detection of electroactive or ion species as well as for decoupling two electrical fields in a capillary electrophoresis column and an electrochemical detector.